Tumor vascularization in many instances is elicited by a tumor-produced angiogenesis factor. In this proposal in vitro studies are outlined concerning the effects of this factor on its apparent target, endothelial cells. Short term cultures of these cells to be isolated from (fetal) calf aorta and vena cava and rabbit aorta will be used to develop a straightforward assay for detecting the presence of the factor by means of its ability to stimulate cell growth (as measured by incorporation of radiolabeled precursors into endothelial cell macro-molecules). This assay will permit isolation and purification of the factor as well as examination of the mechanism(s) of action for the tumor mitogen. Studies at the molecular level on angiogenesis factor (to be isolated most probably from the Walker 256 carcinoma and purified to homogeneity according to various criteria) will center on a) confirming its reported complex composition, b) identifying all components essential for the mitogenic function, c) possibly identifying essential amino acid residues in the protein component (selectively modified by chemical or enzymatic means), and d) seeking alternative, diagnostic (mainly enzymatic) properties in pure factor. The experimental focus of the studies on the mechanism(s) of action will be on first determining whether the factor acts directly upon endothelial cells or upon other blood or vascular components essential for the display of a mitogenic effect. The relationship between these effects and endothelial cell membrane interactions with the factor(s) would be examined next. Structurally modified factor(s) or potentially inhibitory analogs will be used for these studies as well as endothelial cells with selectively modified plasma membranes or with chemically-altered endocytotic properties. These proposed investigations are directed toward making the relationship between the angiogenesis factor and endothelial cells more amenable to biochemical study and toward probing the structure-function relations of the factor(s). The results of such studies should contribute significant insights for devising therapeutic approaches toward regulating tumor angiogenesis factor levels and/or activity and consequently controlling tumor size and metastasis.